Category Archives: RC2

Repeating Crumley et al (1950)

Koch asked me in a comment to replicate an experiment done in 1950 by Helen A Crumley et al demonstrating Tobacco seed growth in deuterium oxide (D2O). The experiment is rather simple (and the figure from the paper is shown below) as Crumley placed 100 seeds in differing amounts of D2O (double distilled water, 33%, 66%, and 99.8% D2O) and analyzed the growth.

Influence of ordinary water, 33%, 66%, and 99% deuterium oxide on tobacco seed germination. Counts made at daily intervals for 39 days. (From Crumley, Fig 3)

So here I am planning the experiment. I will change some things from their experiment. First they placed the seeds on wet cloths (paper towels?), and I will submerge the seeds in the water amounts they used. They also used a variety of plant species (tobacco, clover, radish, Kentucky bluegrass), where I will just use tobacco seeds (but I will try two different species). Finally they talk about their results in terms of percent germination, but it isn’t clear from the paper if they mean number of plants that have exhibited germination, or if they are referring to some amount of growth exhibited by each plant. I will look for both possibilities and report the results as I find them.

In a preliminary experiment I will submerge the plants in water in petri dishes and seal it with parafilm. I will be looking into a more airtight solution as time goes on. I also won’t do 100 seeds but probably on the order of 33 seeds per sample. And in the future I will look into figuring out a way to measure the seed growth.

Original Crumley paper can be found here.

Typical Specimen for the Repeating Crumley Experiment

typical examples, of no germination, beginning germination, obvious germination, etc

Bill Hooker suggested (and rightly so!) that I document what I consider to be typical specimens. I found, today that the zoom feature on the webcam I’m using is quite sufficient for getting close enough to demonstrate this and so I snapped a picture and tried to document. Powerpoint messed me up a little bit, but this is good enough.

The orange box is highlighting a seed that is showing no signs of germination. From my studies I’ve found that the the seed coat becomes slightly transparent just before radicle (the pre root) penetration and you can see the precursor to puncture. The seed coat usually gets lighter too and in this case is really dark in color.

The pink box (hehe) is what I would count as germination when I’m counting the seeds. I see the tip of the radicle penetrating the seed coat and so germination is officially underway.

In the very near dead center of the image is a seed that has what I referred to in an earlier post as an extended radicle. Basically I meant that the radicle was pushed through and the root is now forming. The root will continue to grow until the seed coat has come off, which over to the right it has (there is a green leaf that is cropped on the edge of the image).

The end.

Repeating Crumley Try 2: Day 0

I’m going to start naming this series RC2: Day X so it’s a little shorter for me to type out. Notice there are two new samples to keep track of. I’m doing percentages of D2O in both DI water and deuterium depleted water.

Post about setup is coming soon and so will the live results page.

RC2: Day 1

RC2: Day 2

Looks like there are a few sprouts already…

RC2: Day 3

It should be noted that I shifted all the seeds around in most of the samples so there was a decent distribution in the center of the sample as opposed to being near the edges and near the meniscus. Hopefully everything will remain like this and I won’t have to ever touch the samples for the next 10 days.

RC2: Day 4

I haven’t had much time to study these pictures, but from quick observations it looks like the seeds in 33% d2o mixed with ddw are sprouting before the seeds in 33% d2o mixed with di water. Another interesting thing to note is the seeds in pure di water are sprouting slowly. I’m hoping that my use of nail polish to seal the analyslides hasn’t ruined the experiment.

RC2: Day 5

Repeating Crumley Try 2: Setup

I am on Trial 2 of the Repeating Crumley Experiment. Trial 1 was a pretty decent success, but I ran into some issues regarding image acquisition, experiment stability, and water evaporation. I made some adjustments and began the experiment again. Here is how I setup the second trial:

  • There are 20 seeds of the Dark Virginia variety from the Tobacco Seed Company in each sample.
  • There are 8 water samples: di water, deuterium depleted water, 33% d2o in di water, 66% d2o in di water, 99.9% d2o, 33% d2o in ddw, 66% d2o in ddw, and a sample of di water without any seeds.
  • The sample of di water with no seeds is acting as a control to monitor the possibility of fungal/mold growth. 20 seeds were added to water and allowed to incubate in solution for 30 minutes. At the end of the incubation period the seeds were removed and the sample was sealed.
  • The seeds were added to each sample container (analyslides), and a water type was added immediately after seed addition.
  • After closing the samples, clear nail polish was added around the rim to seal the chambers from the surrounding environment.
  • Data is taken using a Logitech HD Pro Webcam C910.
In the previous trial, the images were taken from below the samples. This resulted in several sample drops. In this trial, I have placed the samples flat on the lab top and am taking data from above the samples. In this manner, the samples have been significantly less agitated then before.
My biggest worry is that the nail polish has/will contaminate(d) the water samples, producing undesirable results. I have ordered a bunch of supplies to test what may create the best seal, while creating the least amount of interaction with the sample water. That experiment will be started as soon as possible and the result will be used to create a third trial run.

RC2: Day 6